Water Exchange Filter (WEX Filter) for Nuclear Magnetic Resonance Studies of Macromolecules

Mori, Susumu; Johnson, M. O'Neil; Berg, Jeremy M.; Zijl, Peter C.M. van

doi:10.1021/ja00105a044

Cited by 56 publications

(59 citation statements)

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“…Data was processed using the Felix 2.1 Software (BioSym) and peak intensities were measured as volume integrals using the Felix 2.1 peak picking routine except for partially overlapped peaks which were measured by frequency domain line fitting using a slice at maximum intensity. The dependencies of the peak intensities on mixing time were fitted to Equation 1 by nonlinear two-parameter fitting (k obs and k obs 1 R 1A ) 19,26,27 ;…”

Section: Methodsmentioning

confidence: 99%

Measurement of water–amide proton exchange rates in the denatured state of staphylococcal nuclease by a magnetization transfer technique

1997

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The rates of hydrogen exchange were measured in a "physiological" denatured state of staphylococcal nuclease using a NMR magnetization transfer experiment suitable for the measurement of exchange rates faster than 0.5 s-1. The results are compared with predicted exchange rates (kex) for the random coil state (Bai et al., Proteins 17:75-86, 1993). No protection factors (= predicted rate/measured rate) larger than 2.4 were observed, consistent with other NMR data which strongly suggest only small amounts of residual secondary structure in this denatured state. Systematically low protection factors (0.51 +/- 0.23) were found for Asp and Glu residues, while high protection factors were observed for Gly (1.60 +/- 0.60). We conclude that the predicted exchange rates (kex) may have an uncertainty of 2- to 3-fold. Thus, for denatured proteins only protection factors with a value of 5 or larger can be assigned structural significance. These results also demonstrate that multidimensional magnetization transfer NMR techniques are powerful tools in this research field due to its ability to measure rapidly exchanging protons (> 05 s-1) with high accuracy.

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Section: Methodsmentioning

confidence: 99%

Measurement of water–amide proton exchange rates in the denatured state of staphylococcal nuclease by a magnetization transfer technique

1997

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“…Slower exchange yields no observable transfer whereas faster exchange broadens the amide resonance beyond detection. Magnetization transfer experiments have been used to monitor protein hydrogen exchange for solvent-exposed positions that exchange in this time regime near neutral pH (14)(15)(16). However, this technique has Abbreviation: CLEANEX-PM, clean chemical exchange-phase modulated.…”

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confidence: 99%

Millisecond time scale conformational flexibility in a hyperthermophile protein at ambient temperature

Hernández

Jenney

Adams

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

164

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Rubredoxin from the hyperthermophile Pyrococcus furiosus is the most thermostable protein characterized to date with an estimated global unfolding rate of 10 ؊6 s ؊1 at 100°C. In marked contrast to these slow global dynamics, hydrogen exchange experiments here demonstrate that conformational opening for solvent access occurs in the Ϸmillisecond time frame or faster at 28°C for all amide positions. Under these conditions all backbone amides with exchange protection factors between 10 4 and 10 6 , for which EX2 exchange kinetics were directly verified, have exchange activation energy values within 2-3 kcal͞mol of that observed for unstructured peptides. The conformational flexibility of this protein is thus sufficient for water and base catalyst access to the exchanging amide with quite limited structural disruption. The common hypothesis that enhanced conformational rigidity in the folded native state underlies the increased thermal stability of hyperthermophile proteins is not supported by these data.

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“…Exchange processes can be generally monitored by dissolving a protein in D 2 O and following the disappearance of amide proton signals in H-N correlation experiments. For exchange processes that are too fast to be determined with this approach, many NMR spectroscopy experiments have been proposed, based on inversion recovery, saturation recovery, WEX, CLEANEX and other experimental schemes, [36][37][38][39][40][41][42] in which the water polarization is selectively manipulated.…”

Section: Characterization Of Intrinsically Disordered Proteinsmentioning

confidence: 99%

High‐Resolution Characterization of Intrinsic Disorder in Proteins: Expanding the Suite of ¹³C‐Detected NMR Spectroscopy Experiments to Determine Key Observables

Bertini¹,

Felli²,

Gonnelli³

et al. 2011

ChemBioChem

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Order in disorder: The characterization of intrinsically disordered proteins by NMR spectroscopy is a necessity on the one hand and a continuous challenge on the other. We propose two experiments that provide diagnostic parameters to monitor the degree of unfolding of a polypeptide. The test was performed on the yeast Cox17 protein, known to gain its function through maturation from an intrinsically disordered state (see figure).

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Water Exchange Filter (WEX Filter) for Nuclear Magnetic Resonance Studies of Macromolecules

Cited by 56 publications

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Measurement of water–amide proton exchange rates in the denatured state of staphylococcal nuclease by a magnetization transfer technique

Measurement of water–amide proton exchange rates in the denatured state of staphylococcal nuclease by a magnetization transfer technique

Millisecond time scale conformational flexibility in a hyperthermophile protein at ambient temperature

High‐Resolution Characterization of Intrinsic Disorder in Proteins: Expanding the Suite of ¹³C‐Detected NMR Spectroscopy Experiments to Determine Key Observables

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Water Exchange Filter (WEX Filter) for Nuclear Magnetic Resonance Studies of Macromolecules

Cited by 56 publications

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Measurement of water–amide proton exchange rates in the denatured state of staphylococcal nuclease by a magnetization transfer technique

Measurement of water–amide proton exchange rates in the denatured state of staphylococcal nuclease by a magnetization transfer technique

Millisecond time scale conformational flexibility in a hyperthermophile protein at ambient temperature

High‐Resolution Characterization of Intrinsic Disorder in Proteins: Expanding the Suite of 13C‐Detected NMR Spectroscopy Experiments to Determine Key Observables

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High‐Resolution Characterization of Intrinsic Disorder in Proteins: Expanding the Suite of ¹³C‐Detected NMR Spectroscopy Experiments to Determine Key Observables